Our overall aim is to better understand how estrogenic hormones regulate their target tissues. We propose to focus our attention on three aspects of estrogen regulation: (1) regulation of estrogen receptor concentrations in cell cultures of target cells, (2) modification of chromatin structure as a primary effect of estrogens, and (3) changes in DNA synthesis, particularly and refractory state, induced by estrogens. Primary cell cultures of uterine and pituitary cells will be used to study the replenishment of estrogen receptors which occurs following receptor translocation to the nucleus and the subsequent dissociation of the estrogen from its receptor. We are interested in whether estrogen receptor replenishment is due to recycling or resynthesis. We will also study whether "processing" (the loss of receptor that has been reported to occur during the binding and translocation process) is an obligatory precess in the action of estrogens. Chromatin studies are based on the theory that the rapid effect of estrogen on transcription of specific genes is modulated by estrogen-receptor effects on chromatin structure. Attempts to assess overall chromatin structure changes by looking for changes in accessability of DNA to various agents or by assaying the degree of supercoiling or chromatin condensation will be attempted. Covalent modification of chromatin proteins will also be assessed. Attempts will be made to develop primary cell culture systems in which estrogen regulation of DNA synthesis can be studied. This will permit the evaluation of the estromedin hypothesis and other theories of growth regulation. We will also attempt to extend our earlier studies on isolated nuclei to determine by reconstitution studies the nature of the factors involved in the estrogen stimulaton of DNA synthesis as well as the induction of the refractory state.